The box a domain of high mobility group box-1 protein as an efficient siRNA carrier with anti-inflammatory effects

• Published in: Journal of cellular biochemistry 2012-01, Vol.113 (1), p.122-131
• Main Authors: Lee, Sanghyun, Song, Hojung, Kim, Hyun Ah, Oh, Binna, Lee, Dong Yun, Lee, Minhyung
• Format: Article
• Language: English
• Subjects: Animals; Anti-inflammation; Cell Line; Cytokines; DNA-Binding Proteins; Gene Expression Regulation; Gene therapy; Genetic Therapy - methods; HEK293 Cells; High mobility group box-1; HMGB Proteins - genetics; HMGB Proteins - metabolism; Humans; Inflammation - immunology; Inflammation - metabolism; Inflammation Mediators - metabolism; Lipopolysaccharides - pharmacology; Macrophages - drug effects; Macrophages - immunology; Mice; NF-kappa B - metabolism; Recombinant peptide; RNA Interference; RNA, Small Interfering - metabolism; siRNA delivery; Tumor Necrosis Factor-alpha - metabolism
• ISSN: 0730-2312; 1097-4644
• DOI: 10.1002/jcb.23334

High mobility group box‐1 (HMGB‐1) is a DNA binding nuclear protein and pro‐inflammatory cytokine. The box A domain of HMGB‐1 (rHMGB‐1A) exerts an anti‐inflammatory effect, inhibiting wild‐type HMGB‐1 (wtHMGB‐1). In this study, HMGB‐1A was evaluated as an siRNA carrier with anti‐inflammatory effects. HMGB‐1A was expressed and purified by consecutive nickel chelate chromatography, cationic exchange chromatography, and polymixin B chromatography. Purified rHMGB‐1A demonstrated an anti‐inflammatory effect, reducing tumor necrosis factor‐α (TNF‐α) in wtHMGB‐1 or lipopolysaccharide (LPS) activated macrophages. In gel retardation assay, rHMGB‐1A formed a stable complex with siRNA at or above a 1:2 weight ratio (siRNA:rHMGB‐1A). A heparin competition assay showed that an siRNA/rHMGB‐1A complex released siRNA more easily than an siRNA/polyethylenimine (PEI, 25 kDa) complex. Luciferase siRNA/rHMGB‐1A reduced firefly luciferase expression at a similar level as luciferase siRNA/PEI complex. Furthermore, TNF‐α siRNA/rHMGB‐1A synergistically reduced TNF‐α expression in LPS activated macrophages. Therefore, rHMGB‐1A may be useful as an siRNA carrier with anti‐inflammatory effects in siRNA therapy for various inflammatory diseases. J. Cell. Biochem. 113: 122–131, 2012. © 2011 Wiley Periodicals, Inc.